1. Field of the Invention
This invention relates to analytical methods, and particularly to analytical identification methods applicable to identification of woods-grown ginseng and mountain-cultivated ginseng.
2. Description of the Related Art
Radix ginseng is the dried root of a perennial plant Panax ginseng C. A. Meyer (Araliaceae). Panax ginseng C. A. Meyer (oriental ginseng) comes in at least three varieties depending on where and how it is grown: wild ginseng, woods-grown ginseng, and mountain-cultivated ginseng (also known as garden ginseng). All of these varieties derive from the same root source.
Wild ginseng grows without human interference in remote mountains and dense forests under the conditions of natural climate, soil, slope, forest canopy, moisture and associated plants. It is a perennial, rare and endangered plant, listed as the first class of national protection species in the Chinese Red Book, 1st Ed. Wild Panax ginseng is not allowed to be collected and traded in China.
Woods-grown ginseng, seeded artificially, grows in undisturbed natural forest for 15-20 years or longer. The cultivation takes advantage of fertile soil, proper temperature, light and water in a broadleaf forest or a mixed forest of broadleafs and conifers, which imitates the natural growth conditions of wild Panax ginseng. Woods-grown ginseng can be induced to have some of the same characteristics of wild Panax ginseng. Because wild Panax ginseng is increasingly scarce, woods-grown ginseng is the next best thing and is universally sought-after for its medicinal properties.
Mountain-cultivated ginseng is planted in a suitable, hilly area after the area has been logged. Mountain-cultivated ginseng is typically picked after 5-6 years of growth.
The length of cultivation and growth conditions determine ginseng characteristics. Wild ginseng is better in terms of its medicinal properties than wood-grown ginseng, which is in turn better than mountain-cultivated ginseng. However, differentiation of ginseng varieties is difficult based on their exterior characteristics or the characteristics of ground ginseng powder.
The exterior characteristics of ginseng root include the rhizome, the main root, the lateral root, rootlets, and annular marks, which may be used to identify a variety of ginseng. Often, however, exterior characteristics don't reflect the internal quality of ginseng accurately and overemphasis of the usefulness of exterior characteristics in the identification process leads to loss of the consumer confidence.
Chemical characterization work on ginseng has begun as early as in 1939 with a large number of reports appearing in the 1960s. The chemical composition of ginseng extracts is very complicated, with about 200 different compounds having been identified. They are divided into several general classes according to their chemical structures:                1) Terpenes: including monoterpenes, such as α-pinene, β-pinene, dihydrocarvone, carvone, etc; sesquiterpenes, such as straight chain sesquiterpenes nerolidol, cis-farnesene, trans-β-farnesene, etc, single ring sesquiterpenes, such as β-bisabolene, α-elemene, β-elemene, etc; bicyclic sesquiterpenes, such as α-selinene, β-selinene, β-caryophyllene, β-santalol, etc.        2) Aliphatic compounds, including alkanes, alkenes, alkynes, alcohols, ethers, aldehydes, ketones, acids, esters etc.        3) Aromatic compounds, such as ethylbenzene, tert-butylbenzene, 1,2-dimethyl benzene, naphthalene, naphthylamine, etc.; and        4) Heterocyclic compounds, such as 2-pentylfuran, 5-methyl-2-furfural, benzofuran, teramethyl pyrazine, 1,2-dimethyl anthraquinone, etc.        
The amount and contents of various compounds in fresh ginseng extracts vary depending on the length of ginseng growth period and cultivation area, which makes it possible to identify different varieties of ginseng by chromatographic analysis.
The concept of chromatographic fingerprint analysis of complex mixtures is known. By comparing chromatographic fingerprints, samples can be identified. Chromatographic fingerprint analysis involves comparing the similarities and differences (relative intensities) of the corresponding peaks of each chromatogram using a quantization process. Chromatographic fingerprint analysis comprises the following steps: (1). Digitization of the fingerprint (peak measurement, e.g. peak area, peak height etc.); and (2) Calculation of fingerprint spectrum similarity.
In the early research on chromatogram fingerprints, the digitization was realized using the peak height or peak area extracted from the main peak of a chromatogram. However, this method used part of the chromatogram characteristics to express the whole fingerprint, discarding certain information and at last reducing the identification sensitivity of chromatogram fingerprint.
Spectra detected by the modern chromatography are stored as a group of binary numbers in the computer. This group of numbers is called a vector, which conveys the holographic information of the entire spectrum. The similarity measured by this kind of vector reflects the similarity of the entire chromatographic fingerprint of the chemical species. Any chemical species can be expressed by a vector. The resemblance between two chemical species can be measured by the similarity between their vectors.
Chromatogram fingerprint similarity can be calculated from one, several, or all peaks in the chromatogram fingerprint. The latter fingerprint similarity is usually used to measure the overall fluctuation of chemical composition in crude drugs.
Spectra obtained by modern chromatography are usually stored as a group of binary numbers, i.e., a vector. A similarity coefficient S between two chromatographic spectra can be calculated by the following formula:
  S  =                    ∑                  i          =          1                n            ⁢                        (                                    x              i                        -                          x              _                                )                ⁢                  (                                    y              i                        -                          y              _                                )                                                          ∑                          i              =              1                        n                    ⁢                                    (                                                x                  i                                -                                  x                  _                                            )                        2                              ⁢                                    ∑                          i              =              1                        n                    ⁢                                    (                                                y                  i                                -                                  y                  _                                            )                        2                              
In this formula, x and y respectively represent the mean values of two groups of chromatographic numbers and xi and yi represent the corresponding points of chromatographic numbers. The obtained related coefficient is Sε[0,1]. Thereby, a similarity between the two groups of numbers can be determined. The similarity gained by this method can reflect the resemblance of the entire spectra of two samples.
Gas chromatography (GC) uses gas as the mobile phase. It separates components according to their partition ratios in the immobile phase and gas-carrying phase, which has the advantages of high resolution, good selectivity, rapid measurement and wide applicability. Accordingly, GC is particularly useful in determining relative composition of plant extracts and other volatile mixtures.
GC-MS is a technique which combines mass spectroscopy with gas chromatography. This coupling technique records mass spectroscopic data of various compounds of mixture after they have been resolved by GC. This allows for an easier identification of each peak in a chromatographic fingerprint.